The long-term goal of this proposal is to understand the control of mtDNA copy number in mammals and hence regulation of intracellular mitochondrial activity and patterns of mitochondrial gene transmission. Using tissue from an evolutionarily related set of artiodactyl species, our approach will focus on three molecular levels of mitochondrial function. (1) Light strand (L-strand) transcripts prime heavy-strand (H-strand) DNA synthesis. the initial event in mtdNA replication. Therefore the role which sequences upstream of L-strand transcript 5' ends play in determining the accuracy and efficiency of transcription will be determined. (2) H-strand synthesis initiates downstream of the L- strand promoter and usually terminates abortively near sequences will conserved in all mammals, thereby creating triple-stranded D- loop DNA. Therefore cis and trans-elements which regulate initiation and termination of D-loop DNA synthesis will be determined. (3) The generation and segregation of heteroplasmic mitochondria in mammalian tissue are related to developmental changes in mtDNA copy number. Therefore the developmental and generation-to-generation fate of heteroplasmic mtDNA in a unique maternal lineage of Holstein cows will be measured to provide a link between the control of mtDNA replication and mitochondrial transmission genetics.